Synthetic Dicer-Substrate siRNAs as Triggers of RNA Interference - RNA Interference from Biology to Therapeutics - page 47

that intervention of TNF-a production is a valid therapeutic approach to prevent the
development of RIF during antitumor therapy.
Two groups have employed the cationic lipid iFECT™ to deliver DsiRNA into
the CNS via IT injection. The BBB presents an obstacle for delivery of large mol-
ecule compounds to the CNS using IV injection; direct injection into the CNS is the
most direct solution to this problem. Dore-Savard and colleagues used anti- Ntsr2
DsiRNAs to study the function of neurotensin receptor 2 (a GPCR) in thermal noci-
ception in rats [ 79 ]. Ntsr2 mediates analgesia, and administration of the synthetic
neurotensin-2 agonist JMV-431 reduces perception of pain. Downregulation of the
Ntsr2 receptor by anti- Ntsr2 DsiRNAs should block the expected analgesic effects
following JMV-431 administration. The anti-Ntsr2 DsiRNAs were given as two IT
injections at spinal levels L5/6 1 day apart using a dose of 1 mg in iFECT™
(0.005 mg/kg). The neurotensin agonist JMV-431 was administered daily at days
1-4 following the DsiRNA injections, and the rats were studied for nociceptive
behavior using the thermal tail-flick test. Analgesic effects from the JMV-431 were
absent on days 1 and 2 and slowly returned to baseline over the next several days.
Reductions in both Ntsr2 mRNA and protein were observed, consistent with an
RNAi-mediated suppression of Ntsr2 gene expression.
LaCroix-Fralish and colleagues studied function of the b3 subunit of the Na + -
K + -ATPase pump ( Atp1b3 ) in the pain response to formalin footpad burns in mice
[ 80 ]. The mouse strain C57BL/6 shows a higher pain response in this test compared
to the A/J strain. Traditional genetic approaches and QTL analysis had previously
implicated the Atp1b3 gene as possibly being involved in this interstrain variability;
however, no role for this gene in nociception had ever been demonstrated. Anti-
Atp1b3 DsiRNAs formulated in iFECT™ were given by IT injection at a dose of
0.5 mg (0.025 mg/kg) once a day for 3 days, and the mice were, thereafter, studied
for pain responses to formalin footpad injection. Mice given the anti- Atp1b3
DsiRNA showed similar pain responses between strains. Within the control DsiRNA
cohort, the C57BL/6 mice showed higher sensitivity than the A/J mice (i.e., a wild-
type response was seen). These results confirmed a role for Atp1b3 in nociception
in the mouse spinal cord.
Sato and colleagues used symmetric 27/27-nt Dicer-substrate siRNAs (with a
2-nt 3¢-overhang on both ends) to treat hepatic cirrhosis in rats using a targeted
liposomal delivery system [ 81 ]. It is thought that collagen production by the hepatic
stellate cells is crucial to development of hepatic cirrhosis following acute or chronic
injury. Targeted liposomes were made using the Lipotrust lipid reagent system con-
jugated to vitamin A as a means to facilitate delivery to the stellate cells. Symmetric
Dicer-substrate siRNAs targeting the collagen chaperone heat shock protein 47 gene
( Serpinh1 , gp46) were administered IV at doses up to 0.75 mg/kg three times
weekly. This treatment, in contrast to control reagents, reduced collagen production
by the stellate cells and prevented fibrosis in several different models of acute cir-
rhosis, including bile duct ligation and chemical injury with dimethylnitrosamine or
carbon tetrachloride.
Kortylewski and colleagues described use of a novel method to improve delivery
of DsiRNAs to cells expressing TLR9 known to bind “CpG-motif” DNA (an unm-
ethylated cytosine-guanine dinucleotide) [ 82 ]. TLR9 is expressed by some immune